Molecular Pharmacology, Vol 18, 215-223, Copyright © 1980 by the American Society for Pharmacology and Experimental Therapeutics

Receptor-Bound Estrogens and Their Metabolites in the Nucleus of the Isolated Rat Liver Parenchymal Cell

¹ Department of Pharmacology and Reproductive Biology Section, Department of Obstetrics and Gynecology, Yale University School of Medicine, 333 Cedar Street, New Haven, Connecticut 06510

Recent studies have suggested that an estrogen receptor is present in rat liver and that hepatic metabolism may modulate receptor binding by estrogens. The metabolism of estrogens and the identity of receptor-bound estrogens were evaluated in the present study. Isolated hepatic parenchymal cells from rats were incubated with 10^-7 M [³H]-17,-estradiol or [³H]-17,-ethinylestradiol for 5 min. [³H]-17,-estradiol was metabolized much more extensively than [³H]-17,-ethinylestradiol in cellular incubation mixtures. A variety of metabolites of each estrogen was detected. In incubations with [³H]-17,-estradiol, estrone was the principle nonpolar radioactivity. In incubations with [³H]-l7,-ethinylestradiol, 2-hydroxy-17,-estradiol was identified as a metabolite. The identity of the receptor-bound [³H]estrogen was then studied after exposure of cells to [³H]-17,-ethinylestradiol or [³H]-17,-estradiol. After incubation of male or female liver cells with [³H]-17,-ethinylestradiol, the cells were homogenized and purified nuclei were prepared. Unchanged 17,-ethinylestradiol and 2-hydroxy-17,-ethinylestradiol were identified as the receptor-bound estrogens. (17,-ethinylestradiol and 2-hydroxy-17,-ethinylestradiol were also capable of binding to the cytosol receptor in vitro.) After incubation of liver cells with [³H]-17,-estradiol, the principle receptor-bound estrogen in the nucleus from female liver cells was unchanged 17,-estradiol, in contrast to the radioactivity in the total cellular incubations, where estrone predominated. The principle nuclear-bound estrogen in male liver was an unidentified metabolite with the chromatographic mobility of 2-hydroxy-17,-estradiol (however, it could not be methylated with catechol-O-methyl transferase). [³H]-17,-estradiol was apparently less effective than [³H]-17,-ethinylestradiol in binding to cellular receptors because of its more extensive metabolism, primarily to estrone. Although estrone binds cytosol receptors in vitro, it is much less effective than 17,-estradiol in the promotion of receptor translocation in intact liver cells. Hepatic metabolism may modulate estrogenic potency in the liver and generate unusual estrogen metabolites which may interact with receptors. In cellular incubations, 17,-ethinylestradiol and its metabolite 2-hydroxy-17,-ethinylestradiol were more effective than the lower amount of unmetabolized 17,-estradiol and the much higher levels of its metabolite estrone in the promotion of nuclear translocation of the estrogen receptor. Estrogen metabolism may contribute to the low receptor interaction and low biologic potency of 17,-estradiol in the liver.

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ACKNOWLEDGMENTS The authors thank Mrs. Gillian Clark for typing the manuscript and Dr. Mitchell J. Weinberger for purification of the rat liver COMT used in this study. The authors also thank Dr. Neil MacLusky for synthesis of some of the catechol estrogens used here and for his helpful advice and Dr. Lawrence Povirk for his helpful advice.